Human mast cells, effectors of immediate hypersensitivity and potentially of innate immunity, have been classified into two phenotypes based on the protease composition of their secretory granules. MCTc cells contain tryptase, chymase, cathepsin G and mast cell carboxypeptidase; MCT cells contain only tryptase. Stem cell factor is the major differentiation factor for human mast cells. The involvement of mast cells in human disease depends on the numbers and types of mast cells recruited, and their activation status and survival. The current proposal addresses fundamental aspects of mast cell biology that relate to these factors. Aim I examines the development and survival of human mast cells. Chymase mRNA expression and potential induction will be examined in MCT cells. The newly-appreciated ability of IL-4 to induce apoptosis in mast cells, and of IL-6 to modulate this process will be explored. New technology allowing mast cells to develop from single cells will be exploited to examine the precursor frequency and proliferative potential of mast cell progenitors. The role of ganglioside GD3 in mast cell development will be examined. Aim 2 explores three potential pathways to modulate activation of human mast cells. GD3 involvement in FcepsilonR1- mediated activation will be determined. Adenosine, known to augment FcepsilonR1-mediated degranulation of human mast cells, is hypothesized to act through an adenosine 2b receptor. FimH, a mannoside-binding component on enterobacterial fimbria, will be examined for its ability to activate human mast cells. Aim 3 will attempt to immortalize human mast cells by forcing mast cell progenitors to express telomerase. This reverse transcriptase maintains telomeres, preserves cell proliferative activity and prevents cell senescence. Phenotypic and functional features of immortalized mast cell lines will be characterized. A better understanding of mast cell development and activation in humans will provide a more precise understanding of their involvement in human disease, and better strategies for therapeutic interventions.